Chronic prostatitis/chronic pelvic pain syndrome (collectively referred to herein as CPPS) is a syndrome of undetermined etiology occurring in men. CPPS is the third of four subgroups of prostatitis recognized by the NIH. Category I encompasses acute bacterial prostatitis, and Category II covers chronic bacterial infection. Category III, CPPS, includes all remaining prostatitis syndromes, and is subdivided into IIIa (inflammatory) and IIIb (non-inflammatory). These sub-categories can be distinguished by the presence of leukocytosis in expressed prostatic secretions or sediment in a post-massage urine sample. Finally, Category IV represents asymptomatic prostatitis, which often is associated with benign prostate hyperplasia.
Prostatitis is extraordinarily common, resulting in approximately 2 million office visits to primary care physicians and urologists in the United States annually [1997 American Urological Association Annual Meeting, National Ambulatory Medical Care Survey, National Center for Health Statistics, 1990 to 1994]. Patients with CPPS suffer from chronic, episodic pain in the perineum or pelvic region, irritative and obstructive voiding symptoms, and adverse effects upon sexual function [Alexander et al., Urology 48:568-574 (1996)]. Men with chronic prostatitis often require repeated physician visits, commonly to different physicians. Medical expenditures relating to CPPD are conservatively estimated to exceed half a billion dollars annually.
Bacterial vs. Non-Bacterial Prostatitis
Given its apparent prevalence, CPPS has defied characterization to an almost astonishing extent. While an enormous number of patients seek the care of a physician because of prostatitis-like symptoms, almost nothing is known about diagnostic criteria, etiology, or objective signs for CPPS. In a survey conducted through the Internet of 163 men with a diagnosis of prostatitis, Alexander and Trissel found that pain in the pelvic region was the most frequently reported and the most severe symptom in such patients [Alexander et al., Urology 48:568-74 (1996)]. It was because of these observations and the paucity of objective criteria for defining the disease, that the National Institute of Diabetes and Digestive and Kidney Diseases working group in prostatitis suggested that the disease be named Chronic Pelvic Pain Syndrome.
One reason for the present state of confusion regarding CPPS is the similarity of CPPS symptoms to the symptoms of bacterial prostatitis. Only about 5 to 10% of patients whose symptoms are consistent with bacterial prostatitis are shown to have infection in the prostate gland [Weidner et al., Infection 19:S109-S190 (1991)]. The misdiagnosis of CPPS as infectious prostatitis, commonly results in unnecessary treatment with multiple courses of antibiotics at enormous cost to patients and to the health care system with no clearly demonstrated benefit to patients.
An enormous amount of effort has attended the search for a fastidious organism as the cause of CPPS. No clear consensus has emerged identifying any such organism as the causative agent. Published studies have proven difficult to interpret due to lack of a standardized definition of CPPS and the variability in methodologies for detecting infectious organisms. Another difficulty with published studies is that the presence of normal flora in the male urethra complicates the interpretation of culture data. Further, many published studies lack control groups to which the findings in men with prostatitis must be compared.
Recently, Krieger et al. reported an extensive study examining trans-perineal prostate biopsies in 135 men with CPPS for bacterial 16s rRNA-encoding DNA sequences [Krieger et al, J. Clin. Microbiol 34:3120-3128 (1996)]. Krieger et al. found that 77% of the CPPS patients had bacterial DNA sequences in their prostatic tissue that were distinct from normal bowel and skin flora. Patients with bacterial DNA sequences in the prostate also had higher numbers of leukocytes in the prostatic fluid compared to patients without such DNA sequences. Krieger et al. concluded that bacteria distinct from the normal bowel and skin flora are present in the prostatic tissue of men with CPPS and are therefore a potential causative agent of CPPS.
In another study by the same group, Berger et. al reported that an aerobic or anaerobic bacterial organism could be cultured from the prostate tissue in 32% of 85 symptomatic men by performing cultures of trans-perineal needle biopsies in a specialized microbiology laboratory [Berger et al., J. Urol. 157:863-865 (1997)].
However, the Krieger et al. and Berger et al. data cannot be interpreted as positively identifying a causative agent for prostatitis because neither study compared the CPPS group with a control group lacking symptoms of CPPS to demonstrate that the prostate tissue of subjects without CPPS symptoms does not contain such bacterial DNA sequences. Furthermore, in preliminary data, the present inventors have detected similar diverse bacterial 16s rRNA-encoding DNA sequences in 8/9 patients undergoing transperineal radioactive seed implantation for localized adenocarcinoma of the prostate, all of whom had no antecedent history of CPPS symptoms [S. Keay et al, Urology 53: 487-491, 1999]. Thus, while the data of Krieger et. al. show that bacterial DNA sequences exist in the prostate of men with CPPS, the presence of these sequences is not sufficient to demonstrate a bacterial origin for CPPS.
What is clear, however, is that some men with CPPS have evidence of inflammation of the prostate. While the cellular and cytokine mediators involved in the inflammatory process have been increasingly clarified in the immunologic literature, few studies have investigated the immunobiology of the prostate gland to determine whether CPPS might be arise from an auto-immune condition.
Autoimmunity and the Prostate
Many diseases are known to result from autoimmunity. The present inventors have discovered that CPPS has an autoimmune component. The inventors hypothesize that either (1) the autoimmune component of CPPS results from an autoimmune attack upon the prostate, or (2) that a chronic inflammatory process is maintained in CPPS patients as a result of a breakdown of immunoregulatory mechanisms in the immediate environment of the prostate.
There is a substantial body of evidence demonstrating the occurrence of immunological activity within the prostate gland. However, the nature and cause of this activity, and whether it is detrimental to the host, has not been determined. Inflammatory infiltrates in the prostate are very common. In one study of 162 cases of surgically resected prostatic tissue inflammatory infiltrates were found in 98% [Kohnen et al., J. Urology 121:755-60 (1979)]. The infiltrating cells consist of monocytes and activated T and B lymphocytes [Theyer et al., Lab Invest. 66:96-107 (1992); Steiner et al., J. Urology 151:480-84 (1994)].
A rare form of prostatic inflammation, granulomatous prostatitis, has been characterized, although the etiology of the inflammation is also unknown. One of the major theories about this disease, however, is that it represents an immune reaction against self prostatic proteins induced by infection or manipulation of the gland by previous biopsy or surgical procedure [Stillwell et al., J. Urology 138:320-23 (1987); Dhundee et al., Histopathology 18:435-41 (1991)].
The disease is also observed after instillation of Bacillus Calmette-Guerin (BCG) into the bladder as a treatment for superficial bladder cancer [Bahnson, J. Urology 146:1368-69(1991)].
Recent observations about the existence of subsets of CD4.sup.+ T cells has yielded fundamental information about immune responses in humans. CD4.sup.+ T cells can be separated into subsets based upon the patterns of cytokines they secrete [Mosmann, Ann NY Acad. Sci. 664:89-92 (1992)). CD4.sup.+ T cells that secrete IFN-.gamma. and IL-2 are called T helper 1 (Th1) cells. Th1 cells mediate cellular immunity, such as delayed hypersensitivity responses. CD4.sup.+ T cells that secrete IL-4 and IL-10 are termed T helper 2 (Th2). Th2 cells are associated with antibody production and allergy. Immune responses mediated by Th1 and Th2 cells can be characterized by the local cytokine environment during the developing immune response.
Zisman et al. found IgG anti-PSA antibody titers to be higher in the serum of men with benign prostate hyperplasia (BPH) compared to controls [Zisman et al., J. Urology 154:1052-55 (1995)]. However, of 17 men with chronic prostatitis, Zisman et al. found no difference in mean antibody titer as compared to controls. Zisman et al. speculate that an immunologic mechanism may play a role in the symptomatology of BPH. An alternative explanation is that a Th1 type of response may be occurring in patients with chronic prostatitis/chronic pelvic pain syndrome. In this event, no antibody response would be expected. A further understanding of the local cytokine environment in the prostate will be critical to understanding the nature of the inflammatory response that is occurring in order to more clearly understand the disease.
There remains a major need in the art for an objective means for diagnosing CPPS, to identify the cause or causes of the syndrome and for CPPS treatments that can result in an improvement in symptoms in men with the disease.
Cytokines
Cytokines are small to medium-sized proteins or glycoproteins which mediate potent biological effects on most cell types. [See Mire-Sluis, et al., eds. Cytokines (1998); also see Thompson, Angus, ed. The Cytokine Handbook (1998)] While cytokines were originally identified as key components in inflammatory processes, it is now known that cytokines are involved in many non-inflammatory physiological processes. Cytokines mediate their effects by binding to specific cell surface receptors which are coupled to intracellular signal transduction and second messenger pathways. More than one hundred cytokines have been identified to date. The inventors have identified the following cytokines as having particular importance in the operation of the present invention: IL8, GM-CSF, IL-.beta. and TNF-.alpha..
IL-8 is a member of the chemokine family of cytokines. IL-8 is produced by many cell types. In general, the biological activity of IL-8 results from its ability to activate the CXC chemokine receptors CXCR1 and CXCR2. Among many other functions, IL-8 acts as a neutrophil chemoattractant and activating factor [Matsushima et al., Cytokine 1:2-13 (1989); Matsushima et al., J. Exp. Med. 167:1883-1893 (1988); Oppenheim et al., Annu. Rev. Immunol. 9:617-648 (1991)]. Synonyms for IL-8 include neutrophil attractant/activating protein (NAP-1), monocyte derived neutrophil activating peptide (MONAP), monocyte derived neutrophil chemotactic factor (MDNCF), neutrophil activating factor (NAF), leukocyte adhesion inhibitor (LAI), Granulocyte chemotactic protein (GCP). IL-8 can be obtained from known commercial sources.
GM-CSF, or granulocytelmacrophage colony stimulating factor, is a survival and growth factor for haematopoietic progenitor cells, a differentiation and activating factor for granulocytic and monocytic cells, and a growth factor for endothelial cells, erythroid cells, megakaryocytes and T cells [Wong et al., Science 228:810-815 (1985); Gough et al., EMBO. J. 4:645-653 (1985); Clarke et al., Science 236: 1229-1237 (1987); Groopman et al., New Eng. J. Med. 321, 1449-1459 (1989)]. GM-CSF is also known as CSFa or pluripoietin-a. GM-CSF can be obtained from known commercial sources.
IL-1.beta. has a wide range of biological activities on many different target cell types including B cells, T cells, and monocytes (Dinarello et al., Adv. Immunol. 44:153-205 (1989); Fuhlbrigge et al., In The Year in Immunology 1988: Immunoregulatory Cytokines and Cell Growth, Vol. 5, Cruse et al., pp.21-37 (1989); di Giovine et al. lmmunol. Today 11:13-20 (1990)]. In vivo, IL-1.beta. induces hypotension, fever, weight loss, neutrophilia, and acute phase response. IL-1.beta. can be obtained from known commercial sources.
IL-6 is a multifunctional cytokine secreted by both lymphoid and non-lymphoid cells which regulates B and T cell function, haematopoiesis and acute phase reactions [Hirano, in The Cytokine Handbook, Thomson, ed., Academic Press, London, pp. 169-190 (1991); Kishimoto, Blood 74:1-10 (1989); Kishimoto, et al., Science 258:593-597 (1992)]. Synonyms for IL-6 include interferon-.beta..sub.2, 26-kDa protein, B cell stimulatory factor 2 (BSF-2), hybridoma/plasmacytoma growth factor (HPGF or IL-HP1), hepatocyte stimulating factor (HSF), monocyte granulocyte inducer type 2 (MGI-2), cytotoxic T cell differentiation and thrombopoietin. IL-6 is produced by a large member of cell types, including lymphoid cells (T cells, B cells) and many non-lymphoid cells including macrophages, bone marrow stromal cells, fibroblasts, keratinocytes, mesangium cells, astrocytes and endothelial cells. IL-6 can be obtained from known commercial sources.
TNF-.alpha. is a potent paracrine and endocrine mediator of inflammatory and immune functions. It is also known to regulate growth and differentiation of a wide variety of cell types. TNF-.alpha. is selectively cytotoxic for many transformed cells, especially in combination with IFN-.gamma.. In vivo, it leads to necrosis of methylcholanthrene-induced murine sarcomas. Many of the actions of TNF-.alpha. occur in combination with other cytokines as part of the "cytokine network" [Manogue et al. In The Cytokine Handbook, Thomson ed., Academic Press, London, p. 241-256 (1991); Fiers, FEBS 285:241-256 (1991); Ruddle, Curr. Opin. Immunol. 4: 327-332 (1992)]. TNF-.alpha. is expressed as a type 11 membrane protein attached by a signal anchor transmembrane domain in the propeptide and is processed by a matrix metalloproteinase (Gearing et al., Nature 338:225-228 (1994). TNF-.alpha. can be obtained from known commercial sources.